Cellular differentiation is at a basic level a specification of the global protein complement, or proteome. The scale and speed of these changes are remarkable in the case of terminally differentiated cells such as erythrocytes, spermatozoa, and others. Global remodeling of the proteome consists of the programmed elimination of most generic constituents of the cell in parallel with abundant synthesis of a small number of new, cell-type-specific proteins such as globin. Reticulocytes are a canonical example of a proteome in rapid transition, yet, even in this case, the mechanisms that drive rapid turnover of normally stable proteins largely remain to be identified. We have found that Ube2O/E2-230K is an ubiquitin-conjugating enzyme expressed selectively in the erythroid lineage and that null mutations in its gene have a highly specific phenotype of microcytic anemia. Further analysis of this murine mutant, hem9, suggests the hypothesis that Ube2O is a broad-spectrum ubiquitinating enzyme that plays a key role in the global remodeling of the erythroid proteome. To assess this model, an integrated set of approaches is proposed, incorporating mutant analysis, mass spectrometry, and in vitro biochemistry. A major point of focus will be to identify substrates of Ube2O in an unbiased and global manner. To date there has been significant progress towards this goal, as the globin chaperone AHSP, globin itself, and ribosomes have been provisionally assigned as Ube2O substrates. Accordingly, a distinctive phenotype of hem9 is markedly elevated levels of 80S ribosomes. In contrast, low ribosome levels are the signature feature of Diamond Blackfan anemia. Ube2O was found to be sufficient to induce ribosome degradation by dox-inducible expression in HEK293 cells; no other erythroid-specific factors are needed. The ubiquitination and degradation of AHSP and ribosomes will be reconstituted in vitro and in highly purified systems; the specificity determinants of these reactions will be mapped for both substrate and enzyme. In parallel with these biochemical investigations, the physiological effects of Ube2O loss will be studied at higher resolution through analysis of protein synthesis in the mutant reticulocytes, and bone marrow transplantation will be used to test whether hem9 erythroid phenotypes are cell-autonomous, as predicted by the hypothesis that Ube2O reshapes the proteome. Ultimately, our studies are expected to shed light on the fundamental process of proteome remodeling and provide new insights into underlying mechanisms of both microcytic anemia and Diamond Blackfan anemia.